Binding of beta-secretase to a peptide inhibitor-carrying SAM.

A novel disulfide, which carried a specific inhibitor for beta-secretase (KMI360) at both ends, was prepared by the coupling of 11,11'-dithiobisundecanoic acid (DTUA) with the inhibitor. The compound obtained (DTUA-KMI360) formed a self-assembled monolayer (SAM) on a gold electrode as proven by cyclic voltammetry (CV) using hydroquinone as a probe. Furthermore, DTUA-KMI360 could be accumulated as a SAM on a gold colloid deposited on a glass plate (Au colloid-glass chip) as proven by both the red-shift and the increase in absorbance of the gold colloid corresponding to localized surface plasmon resonance (LSPR). When the SAM-modified Au colloid-glass chip was immersed in a solution of aspartyl proteases, pepsin and beta-secretase, the absorbance of the chip at 550nm corresponding to LSPR of the gold colloid further increased and was slightly red-shifted, whereas coexistence of a free inhibitor obstructed these phenomena. Adsorption of the enzymes was promoted by the incorporation of a zwitterionic group into the SAM, while non-specific adsorption to the mixed SAM was significantly reduced. The optimal ratio of omega-zwitterionic alkanethiol, 3-[(6-mercaptohexyl)-N,N-dimethylamino]propane-1-sulfonic acid (C(6)-SPB), and DTUA-KMI360 in the SAM for the binding of enzymes was found to be DTUA-KMI360:C(6)-SPB=1:11 using polarization modulation infrared reflection absorption spectroscopy (PM-IR-RAS). From increasing profiles of absorbance of the Au colloid-glass chip, the association constant (K(assoc)) for pepsin with the inhibitor on the SAM was determined, whereas that for beta-secretase could not be due to the strong binding of the enzyme to the inhibitor, resulting in the absence of the dissociation process. The results suggested that the SAM of the enzyme inhibitor can be used for both investigation of enzymes and removal of target enzymes from biological fluids.

[1]  H. Kitano,et al.  Sensing capabilities of colloidal gold modified with a self-assembled monolayer of a glucose-carrying polymer chain on a glass substrate. , 2004, Langmuir : the ACS journal of surfaces and colloids.

[2]  N. Kanayama,et al.  Substrate Monolayers as Electrochemical Sensing Elements for α-Chymotrypsin , 2002 .

[3]  Kazuhiro Matsuura,et al.  Kinetic study on the binding of lectin to mannose residues in a polymer brush. , 2009, Colloids and surfaces. B, Biointerfaces.

[4]  George M. Whitesides,et al.  Structures of self-assembled monolayer films of organosulfur compounds adsorbed on gold single crystals: electron diffraction studies , 1988 .

[5]  Robert B. Gennis,et al.  Biomembranes: Molecular Structure and Function , 1988 .

[6]  David N. Reinhoudt,et al.  Sensor functionalities in self-assembled monolayers , 2000 .

[7]  D. Reinhoudt,et al.  Molecular Recognition by Self-Assembled Monolayers of Cavitand Receptors , 1994, Science.

[8]  Ashutosh Chilkoti,et al.  A colorimetric gold nanoparticle sensor to interrogate biomolecular interactions in real time on a surface. , 2002, Analytical chemistry.

[9]  R. G. Freeman,et al.  Preparation and Characterization of Au Colloid Monolayers , 1995 .

[10]  S. Kitazume,et al.  KMI-358 and KMI-370, highly potent and small-sized BACE1 inhibitors containing phenylnorstatine. , 2004, Bioorganic & medicinal chemistry letters.

[11]  W. Murphy,et al.  Multifunctional mixed SAMs that promote both cell adhesion and noncovalent DNA immobilization. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[12]  R. Murray,et al.  Monolayer-protected cluster molecules. , 2000, Accounts of chemical research.

[13]  H. Kitano,et al.  Galactose-containing amphiphiles prepared with a lipophilic radical initiator. , 1995, Bioconjugate chemistry.

[14]  Joseph Wang,et al.  Hydrazine Detection Using a Tyrosinase-Based Inhibition Biosensor , 1995 .

[15]  Timothy D. H. Bugg,et al.  Introduction to Enzyme and Coenzyme Chemistry , 1997 .

[16]  Milan Mrksich,et al.  Probing Protein–Carbohydrate Interactions with Microarrays of Synthetic Oligosaccharides , 2004, Chembiochem : a European journal of chemical biology.

[17]  Shuichi Takayama,et al.  Zwitterionic SAMs that Resist Nonspecific Adsorption of Protein from Aqueous Buffer. , 2001, Langmuir : the ACS journal of surfaces and colloids.

[18]  B. Fenderson,et al.  Specific interaction between Lex and Lex determinants. A possible basis for cell recognition in preimplantation embryos and in embryonal carcinoma cells. , 1989, The Journal of biological chemistry.

[19]  Ralph G. Nuzzo,et al.  ADSORPTION OF BIFUNCTIONAL ORGANIC DISULFIDES ON GOLD SURFACES , 1983 .

[20]  Ralph G. Nuzzo,et al.  Spontaneously organized molecular assemblies. 3. Preparation and properties of solution adsorbed monolayers of organic disulfides on gold surfaces , 1987 .

[21]  H. Kitano,et al.  Self-Assembled Monolayers as Novel Biomembrane Mimetics. 1. Characterization of Cytochrome c Bound to Self-Assembled Monolayers on Silver by Surface-Enhanced Resonance Raman Spectroscopy , 1995 .

[22]  G. Rechnitz,et al.  Toxin detection using a tyrosinase-coupled oxygen electrode. , 1993, Analytical chemistry.

[23]  J. Treanor,et al.  Beta-secretase cleavage of Alzheimer's amyloid precursor protein by the transmembrane aspartic protease BACE. , 1999, Science.

[24]  Jean-Marie Lehn,et al.  Comprehensive Supramolecular Chemistry , 1996 .

[25]  H. Endo,et al.  Inclusion of bisphenols by a self-assembled monolayer of thiolated calix[6]arene on a gold surface. , 2005, Environmental science & technology.

[26]  Kitano,et al.  Recognition of Amphiphiles with Many Pendent Galactose Residues by Ricinus communis Agglutinin. , 1999, Journal of colloid and interface science.

[27]  R. Nuzzo,et al.  Synthesis, Structure, and Properties of Model Organic Surfaces , 1992 .

[28]  H. Kitano,et al.  INCLUSIONAL COMPLEXATION BY CYCLODEXTRINS AT THE SURFACE OF SILVER AS EVIDENCED BY SURFACE-ENHANCED RESONANCE RAMAN SPECTROSCOPY , 1995 .

[29]  H. Kitano,et al.  Regio- and Stereoselective Complexation by a Self-Assembled Monolayer of Thiolated Cyclodextrin on a Gold Electrode , 1997 .

[30]  L. Chau,et al.  Colloidal gold-modified optical fiber for chemical and biochemical sensing. , 2003, Analytical chemistry.

[31]  R. Nuzzo,et al.  Molecular ordering of organosulfur compounds on Au(111) and Au(100): Adsorption from solution and in ultrahigh vacuum , 1993 .

[32]  Noo Li Jeon,et al.  Structure and Stability of Patterned Self-Assembled Films of Octadecyltrichlorosilane Formed by Contact Printing , 1997 .

[33]  Charles T. Campbell,et al.  Binding and Dissociation Kinetics of Wild-Type and Mutant Streptavidins on Mixed Biotin-Containing Alkylthiolate Monolayers , 2000 .

[34]  H. Kitano,et al.  Stereoselective inclusion of DOPA derivatives by a self-assembled monolayer of thiolated cyclodextrin on a gold electrode , 1999 .

[35]  G. Whitesides,et al.  Formation of monolayer films by the spontaneous assembly of organic thiols from solution onto gold , 1989 .

[36]  M. Porter,et al.  Reductive desorption of alkanethiolate monolayers at gold: a measure of surface coverage , 1991 .

[37]  Wolfgang Knoll,et al.  MOLECULAR RECOGNITION AT SELF-ASSEMBLED MONOLAYERS - THE CONSTRUCTION OF MULTICOMPONENT MULTILAYERS , 1993 .

[38]  T Kobayashi,et al.  Local plasmon sensor with gold colloid monolayers deposited upon glass substrates. , 2000, Optics letters.

[39]  K. A. Lee.  Electron transfer into self-assembling monolayers on gold electrodes , 1990 .

[40]  H. Kitano,et al.  Recognition of novel lipopolypeptides with many pendent sugar residues by lectin. , 2001, Bioconjugate chemistry.

[41]  George M. Whitesides,et al.  Convenient methods for patterning the adhesion of mammalian cells to surfaces using self-assembled monolayers of alkanethiolates on gold , 1993 .

[42]  H. Kitano,et al.  Recognition of novel amphiphiles with many pendent mannose residues by Con A. , 1999, Bioconjugate chemistry.

[43]  A. Spatola,et al.  An Advantageous Method for the Rapid Removal of Hydrogenolysable Protecting Groups under Ambient Conditions; Synthesis of Leucine-enkephalin , 1980 .

[44]  H. Ringsdorf,et al.  Molecular Architecture and Function of Polymeric Oriented Systems: Models for the Study of Organization, Surface Recognition, and Dynamics of Biomembranes , 1988 .

[45]  S. Romeo,et al.  Plasmepsin II inhibition and antiplasmodial activity of Primaquine-Statine 'double-drugs'. , 2004, Bioorganic & medicinal chemistry letters.

[46]  J. F. Stoddart,et al.  SUPPORTED MONOLAYERS CONTAINING PREFORMED BINDING SITES. SYNTHESIS AND INTERFACIAL BINDING PROPERTIES OF A THIOLATED BETA -CYCLODEXTRIN DERIVATIVE , 1995 .

[47]  D. Frizzell,et al.  New pepstatin analogues: synthesis and pepsin inhibition. , 1991, Journal of medicinal chemistry.

[48]  I. Harada,et al.  Ultraviolet resonance Raman study on the binding mode of enkephalin to phospholipid membranes , 1992 .

[49]  Y. Umezawa,et al.  An SPR-based screening method for agonist selectivity for insulin signaling pathways based on the binding of phosphotyrosine to its specific binding protein. , 2000, Analytical chemistry.

[50]  C. Chidsey,et al.  Chemical functionality in self-assembled monolayers: structural and electrochemical properties , 1990 .

[51]  H. Kitano,et al.  Dehydration Effect on the Recognition of Amphiphiles with Many Pendent Mannose Residues by Concanavalin A , 2001 .

[52]  C. Ruan,et al.  Thionine covalently tethered to multilayer horseradish peroxidase in a self-assembled monolayer as an electron-transfer mediator. , 1998, Analytical chemistry.

[53]  H. Kitano,et al.  Molecular Recognition by Self-Assembled Monolayers of Cyclodextrin on Ag† , 1997 .

[54]  G. Olivecrona,et al.  Interaction of lipoprotein lipase with heparin fragments and with heparan sulfate: stoichiometry, stabilization, and kinetics. , 1996, Biochemistry.

[55]  H. Kitano,et al.  Interfacial Recognition of Sugars by Boronic Acid-Carrying Self-Assembled Monolayer† , 2000 .

[56]  George M. Whitesides,et al.  BIOSPECIFIC ADSORPTION OF CARBONIC ANHYDRASE TO SELF-ASSEMBLED MONOLAYERS OF ALKANETHIOLATES THAT PRESENT BENZENESULFONAMIDE GROUPS ON GOLD , 1995 .

[57]  F. Schreiber.  Structure and growth of self-assembling monolayers , 2000 .